Timms et al. report that two estrogenic chemicals in widespread use--the birth control hormone, ethinylestradiol and the plastic molecule, bisphenol A --cause adverse effects on the development of the prostate in fetal mice, at levels beneath common exposures experienced by the American public. Their findings may provide important insights into factors responsible for increases in prostate disease, including prostate cancer.

Babies in the womb are exposed to ethinylestradiol when women on the pill accidentally become pregnant and continue to take birth control. This happens to some 2 million women in the US and Europe each year.

Exposure to bisphenol A (BPA) takes place because of its widespread use for food and water containers, and because the chemical bonds which hold BPA together in its polymerized form readily degrade, allowing BPA to leach into materials with which it comes into contact.

Their study also confirms impacts of exposure to bisphenol A at levels far lower the EPA's current safety standard. Industry scientists have claimed these results can't be replicated. Recent comments by industry spokespeople have been blatantly wrong.

What did they do? Timms et al. fed corn oil spiked with different contaminants to pregnant mice and then examined the prostate glands of their male offspring at birth (day 19 after fertilization).

They divided their animals into four groups and administered the treatment for four days (days 14-18 after fertilization):

In a separate study they compared the effects of a high dose of DES (200 µg/kg/day) to a control group using the same procedures.

Just prior to birth, the fetuses were removed via Caesarian section. They examined one male fetus per litter, selecting one that had developed in utero between a male and a female. They took this step because prior work has shown that intrauterine position can have significant effects on reproductive tract development. By selecting individuals from the same intrauterine position, Timms et al. eliminated a source of confusion in their data.

Timms et al. dissected out the prostates and used imaging techniques to create a 3-dimensional image of the fetuses developing prostate glands. This allowed very precise analyses of aspects of the glands' shape and size.

They also used immunological techniques to identify cells that were proliferating in different regions of the developing prostate. This allowed them to compare the impact of the treatments on cell division patterns.

What did they find? Development of fetal prostates were significantly altered in the animals exposed to low-level DES, BPA and EE, compared to controls. The effects by BPA and EE were virtually identical to those of DES.

These images show the urogenital sinus of mice just before birth. The red is the urethra. Green, yellow and blue are dorsal, lateral and ventral prostate ducts, respectively. Images shown are from individuals closest to the mean for each treatment group.

Animals treated with DES, BPA and EE all differed significantly from the controls. Total number of ducts were increased as was total volume. For example, BPA increased volume by 81% and duct number by 41%.

In contrast to the increases in prostate duct number and volume by extremely low levels of DES, BPA and EE (above), high level exposure to DES (2000 times higher than the low-level exposure) completely stopped duct formation in the dorsal and lateral prostate. This can be seen in the figures to the left, which are computer reconstructions of the urogenital sinus at day 19, in a different visualization than the figures above.

The prostates of animals in the control group had well developed dorsal and lateral prostate ducts (DP, LP) but these were eliminated in the high DES group, as were the coagulating glands (CG). The ventral ducts (VP) were much smaller with a different growth pattern.

At low levels each of the estrogenic chemicals caused increases in the number of proliferating cells.

Low-level exposures also caused an abnormal narrowing in the urethra at the neck of the bladder.

What does it mean? This study is important for 3 reasons:

It implicates two common human exposures in the development of prostate disease. Millions of fetuses are exposed each year to ethynilestradiol and bisphenol A, at levels above those used in these experiments. Over the past several decades several prostate conditions, including prostate cancer, have increased in frequency. These mouse experiments do not establish causation. Instead they point to a biological process by which exposures of these compounds could be contributing to prostate disease. No human data are currently available to test whether similar events occur in people. As Timm et al. suggest in their paper, however, there are strong biological reasons to believe these results are relevant.

It confirms and extends an earlier report by Nagel et al. that extremely low-dose exposure to bisphenol during fetal development causes adverse effects on prostate development. That study measured prostate weight. In this paper, Timm et al. provide detailed insight into what tissues within the prostate are affected by BPA and these other estrogenic compounds. These results also confirm the validity of DES as a positive control for low dose exposures to estrogenic substances. This had been a point of debate: when industry's use of DES as a positive control failed, they then asserted it wasn't appropriate, instead of acknowledging that their failure meant their experiment had not worked and should be disregarded.

It provides an extremely clear example of the fact that high dose experiments cannot reliably predict possible adverse consequences of low-dose exposures. As Timm et al. show, exposures to relatively high levels of DES shut down prostate gland formation, while exposures to DES at a dose 2000-times lower enhanced duct formation. Regulatory toxicology has been conducted for decades on the assumption that high dose experiments would be sufficient. Clearly that is wrong. Hence these results represent a major challenge to the standard procedures for identifying safe levels of exposure.

These results come directly on the heels of an overview published in Environmental Health Perspectives of the adverse effects that have been tied to bisphenol A through laboratory experiments with animals and cells. That list is long. Many of the effects are caused by levels well within the range to which most Americans are exposed. The authors of that commentary, one of whom (vom Saal) is an an author of this paper, conclude that the current regulatory standard for BPA, established by the EPA in 1988, is out of date. Indeed, more than half of the relevant studies have been published since 2002. They also report a strong bias: none of the 11 industry-funded studies to date report adverse effects, whereas over 90% of the more than 100 studies funded by government find impacts.

This new paper by Timms et al. adds to the weight of evidence, now strong, that BPA standards should be tightened very significantly.